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Characteristics and Resistance to Antimicrobials of Staphylococcus Aureus and Staphylococcus Pseudintermedius Isolated from Pet Animals and Their Owners

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Characteristics and Resistance to Antimicrobials of Staphylococcus Aureus and Staphylococcus Pseudintermedius Isolated from Pet Animals and Their Owners LITHUANIAN UNIVERSITY OF HEALTH SCIENCES VETERINARY ACADEMY Jūratė Stankevičienė CHARACTERISTICS AND RESISTANCE TO ANTIMICROBIALS OF STAPHYLOCOCCUS AUREUS AND STAPHYLOCOCCUS PSEUDINTERMEDIUS ISOLATED FROM PET ANIMALS AND THEIR OWNERS Doctoral Dissertation Agricultural Sciences, Veterinary (A 002) Kaunas, 2019 Dissertation has been prepared at the Department of Veterinary Patho- biology (former Infectious Diseases) of the Veterinary Academy of the Lithuanian University of Health Sciences during the period of 2011–2019. Scientific Supervisor Prof. Dr. Jūratė Šiugždaitė (Lithuanian University of Health Sciences, Agricultural Sciences, Veterinary – A 002). Dissertation is defended at the Veterinary Research Council of the Lithuanian University of Health Sciences: Chairperson Prof. Dr. Alius Pockevičius (Lithuanian University of Health Sciences, Agricultural Sciences, Veterinary – A 002). Members: Assoc. Prof. Dr. Gintaras Daunoras (Lithuanian University of Health Sciences, Agricultural Sciences, Veterinary – A 002); Dr. Marius Virgailis (Lithuanian University of Health Sciences, Agricul- tural Sciences, Veterinary – A 002); Prof. Dr. Algimantas Paulauskas (Vytautas Magnus University, Natural Sciences, Biology – N 010); Dr. Olga Muter (University of Latvia, Natural Sciences, Biology – N 010). Dissertation will be defended at the open session of the Veterinary Research Council of Lithuanian University of Health Sciences, Veterinary Academy, 2019 on 30 of August at 10 p.m. in the Dr. S. Jankauskas auditorium. Address: Tilžės 18, LT-47181, Kaunas, Lithuania. LIETUVOS SVEIKATOS MOKSLŲ UNIVERSITETAS VETERINARIJOS AKADEMIJA Jūratė Stankevičienė STAPHYLOCOCCUS AUREUS IR STAPHYLOCOCCUS PSEUDINTERMEDIUS, IŠSKIRTŲ IŠ GYVŪNŲ AUGINTINIŲ IR JŲ SAVININKŲ, CHARAKTERIZAVIMAS IR ATSPARUMAS ANTIMIKROBINĖMS MEDŽIAGOMS Daktaro disertacija Žemės ūkio mokslai, veterinarija (A 002) Kaunas, 2019 Disertacija rengta 2011–2019 metais Lietuvos sveikatos mokslų universi- tete, Veterinarijos akademijoje, Veterinarinės patobiologijos katedroje (bu- vusi Užkrečiamųjų ligų katedra), Mikrobiologijos laboratorijoje. Mokslinė vadovė prof. dr. Jūratė Šiugždaitė (Lietuvos sveikatos mokslų universitetas, že– mės ūkio mokslai, veterinarija – A 002). Disertacija ginama Lietuvos sveikatos mokslų universiteto Veterinarijos mokslo krypties taryboje: Pirmininkas prof. dr. Alius Pockevičius (Lietuvos sveikatos mokslų universitetas, žemės ūkio mokslai, veterinarija – A 002). Nariai: doc. dr. Gintaras Daunoras (Lietuvos sveikatos mokslų universitetas, žemės ūkio mokslai, veterinarija – A 002); dr. Marius Virgailis (Lietuvos sveikatos mokslų universitetas, žemės ūkio mokslai, veterinarija – A 002); prof. dr. Algimantas Paulauskas (Vytauto Didžiojo universitetas, gamtos mokslai, biologija – N 010); dr. Olga Muter (Latvijos universitetas, gamtos mokslai, biologija – N 010). Disertacija ginama viešame Veterinarijos mokslo krypties tarybos posėdyje 2019 m. rugpjūčio 30 d. 10 val. Lietuvos sveikatos mokslų universiteto Veterinarijos akademijos dr. S. Jankausko auditorijoje. Disertacijos gynimo vietos adresas: Tilžės g. 18, LT-47181, Kaunas, Lietuva. CONTENTS ABBREVIATIONS ....................................................................................... 8 INTRODUCTION ......................................................................................... 9 1. LITERATURE REVIEW ........................................................................ 11 1.1. History .............................................................................................. 11 1.2. Taxonomy......................................................................................... 11 1.3. Colonization of Staphylococcus aureus and Staphylococcus pseudintermedius ..................................................................................... 12 1.3.1. Staphylococcus aureus in canine and feline ........................... 12 1.3.2. Staphylococcus aureus in human ............................................ 13 1.3.3. Staphylococcus pseudintermedius in dogs and cats ................ 13 1.3.4. Staphylococcus pseudintermedius in human .......................... 14 1.4. Staphylococcus aureus and Staphylococcus pseudintermedius as pathogens ............................................................................................ 14 1.4.1. Diseases caused by Staphylococcus aureus and Staphylococcus pseudintermedius in dogs and cats ......................... 14 1.4.2. Diseases caused by Staphylococcus aureus and Staphylococcus pseudintermedius in human .................................... 14 1.5. General characteristics and biochemical properties of Staphylococcus aureus and Staphylococcus pseudintermedius .............. 15 1.6. Virulence factors of staphylococci ................................................... 17 1.7. Identification of staphylococci ......................................................... 20 1.7.1. Phenotypic identification ........................................................ 20 1.7.2. Genotypic identification ......................................................... 22 1.7.3. DNA sequencing ..................................................................... 23 1.8. Resistance to antimicrobials ............................................................. 25 1.8.1. Antimicrobial resistance of Staphylococcus pseudintermedius .............................................................................. 26 1.8.2. Antimicrobial resistance of Staphylococcus aureus ............... 28 1.8.3. Antimicrobial susceptibility testing ........................................ 29 1.9. Interspecies transmission of staphylococci ...................................... 31 2. MATERIAL AND METHODS .............................................................. 33 2.1. Collection of bacterial samples ........................................................ 33 2.2. Isolation of pure culture of the staphylococci .................................. 35 2.3. Phenotypic identification of staphylococci ...................................... 35 2.3.1. Detection of coagulase-positive staphylococci ....................... 35 2.3.2. Determination of virulence factors for Staphylococcus aureus and Staphylococcus pseudintermedius ................................. 35 5 2.3.3. Biochemical characteristics of Staphylococcus aureus and Staphylococcus pseudintermedius .................................................... 36 2.4. Genotypic identification of Staphylococcus aureus and Staphylococcus pseudintermedius ........................................................... 38 2.5. Antimicrobial susceptibility of Staphylococcus aureus and Staphylococcus pseudintermedius ........................................................... 39 2.5.1. Antimicrobial resistance of Staphylococcus aureus and Staphylococcus pseudintermedius detection by disc diffusion method .............................................................................................. 39 2.5.2. The minimal inhibitory and minimum bactericidal concentrations of antimicrobials against Staphylococcus aureus and Staphylococcus pseudintermedius .................................................... 40 2.5.3. Genotypic identification of resistance genes in Staphylococcus aureus and Staphylococcus pseudintermedius ... 41 2.6. Statistical analysis ............................................................................ 42 2.7. Sequencing and phylogenetic analysis ............................................. 43 3. RESULTS ................................................................................................ 45 3.1. Description of population ................................................................. 45 3.2. Isolation of Staphylococcus aureus and Staphylococcus pseudintermedius ..................................................................................... 46 3.2.1. Detection of virulence factors in Staphylococcus aureus and Staphylococcus pseudintermedius ............................................. 47 3.2.2. Biochemical identification of Staphylococcus aureus and Staphylococcus pseudintermedius .................................................... 47 3.2.3. Genotypic identification of Staphylococcus aureus and Staphylococcus pseudintermedius .................................................... 48 3.3. Prevalence of Staphylococcus aureus and Staphylococcus pseudintermedius ..................................................................................... 49 3.3.1. Staphylococcus aureus in canine ............................................ 50 3.3.2. Staphylococcus pseudintermedius in canine ........................... 50 3.3.3. Staphylococcus aureus in feline.............................................. 51 3.3.4. Staphylococcus pseudintermedius in feline ............................ 52 3.3.5. Staphylococcus aureus in humans .......................................... 52 3.3.6. Staphylococcus pseudintermedius in humans ......................... 53 3.4. Phylogenetic analysis of Staphylococcus aureus and Staphylococcus pseudintermedius ........................................................... 55 3.5. Staphylococci genome comparisons ................................................ 56 3.6. Phenotypic antimicrobial susceptibility of Staphylococcus aureus and Staphylococcus pseudintermedius ........................................ 57 6 3.6.1. Resistance of Staphylococcus pseudintermedius in companion animals....................................................................... 58 3.6.2. Resistance of Staphylococcus pseudintermedius
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